1. Field of the Invention
This invention relates to the production of fuel alcohol from cellulose. More specifically, this invention relates to the pretreatment of cellulose feedstocks for ethanol production. The pretreatment reaction of feedstocks chosen with a ratio of arabinan plus xylan to non-starch polysaccharides (AX/NSP) of greater than about 0.39 produces a superior substrate for enzymatic hydrolysis than other feedstocks. These pretreated feedstocks are uniquely suited to ethanol production. Examples of feedstocks that could be chosen in such a pretreatment process include some varieties of oat hulls and corn cobs, and feedstocks selectively bred for high AX/NSP.
2. Brief Description of the Prior Art
The possibility of producing ethanol from cellulose has received much attention due to the availability of large amounts of feedstock, the desirability of avoiding burning or landfilling the materials, and the cleanliness of the ethanol fuel. The advantages of such a process for society are described, for example in a cover story of the ATLANTIC MONTHLY, (April 1996).
The natural cellulosic feedstocks for such a process typically are referred to as "biomass`. Many types of biomass, including wood, agricultural residues, herbaceous crops, and municipal solid wastes, have been considered as feedstocks for ethanol production. These materials primarily consist of cellulose, hemicellulose, and lignin. This invention is concerned with converting the cellulose to ethanol. The familiar corn starch-to-ethanol process, in which the starch is converted to ethanol using sulfurous acid and amylase enzymes, lies outside the scope of this invention.
Cellulose is a polymer of the simple sugar glucose connected by beta 1,4 linkages. Cellulose is very resistant to degradation or depolymerization by acid, enzymes, or micro-organisms. Once the cellulose is converted to glucose, the resulting sugar is easily fermented to ethanol using yeast. The difficult challenge of the process is to convert the cellulose to glucose.
The oldest methods studied to convert cellulose to glucose are based on acid hydrolysis (review by Grethlein, Chemical Breakdown Of Cellulosic Materials, J.APPL.CHEM. BIOTECHNOL. 28:296-308 (1978)). This process can involve the use of concentrated or dilute acids. The concentrated acid process uses 72%, by weight, sulfuric acid or 42%, by weight, hydrochloric acid at room temperature to dissolve the cellulose, followed by dilution to 1% acid and heating to 100.degree. C. to 120.degree. C. for up to three hours to convert cellulose oligomers to glucose monomers. This process produces a high yield of glucose, but the recovery of the acid, the specialized materials of construction required, and the need to minimize water in the system are serious disadvantages of this process. Similar problems are encountered when concentrated organic solvents are used for cellulose conversion.
The dilute acid process uses 0.5% to 2%, by weight, sulfuric acid at 180.degree. C. to 240.degree. C. for several minutes to several hours. BRINK (U.S. Pat. Nos. 5,221,537 and 5,536,325) describes a two-step process for the acid hydrolysis of lignocellulosic material to glucose. The first (mild) step depolymerizes the hemicellulose to xylose and other sugars. The second step depolymerizes the cellulose to glucose. The low levels of acid overcome the need for chemical recovery. However, the maximum glucose yield is only about 55% of the cellulose, and a high degree of production of degradation products can inhibit the fermentation to ethanol by yeast. These problems have prevented the dilute acid hydrolysis process from reaching commercialization.
To overcome the problems of the acid hydrolysis process, cellulose conversion processes have been developed using two steps: (1) a pretreatment, and (2) a treatment comprising enzymatic hydrolysis. The purpose of pretreatment is not to hydrolyze the cellulose completely to glucose but, rather, to break down the integrity of the fiber structure and make the cellulose more accessible to attack by cellulase enzymes in the treatment phase. After a typical pretreatment of this type, the substrate has a muddy texture. Pretreated materials also look somewhat similar to paper pulp, but with shorter fibers and more apparent physical destruction of the feedstock.
The goal of most pretreatment methods is to deliver a sufficient combination of mechanical and chemical action, so as to disrupt the fiber structure and improve the accessibility of the feedstock to cellulase enzymes. Mechanical action typically includes the use of pressure, grinding, milling, agitation, shredding, compression/expansion, or other types of mechanical action. Chemical action typically includes the use of heat (often steam), acid, and solvents. Several known pretreatment devices will be discussed below, and with specific reference to extruders, pressurized vessels, and batch reactors.
A typical treatment by enzymatic hydrolysis is carried out by mixing the substrate and water to achieve a slurry of 5% to 12%, by weight of cellulose, and then adding cellulase enzymes. Typically, the hydrolysis is run for 24 to 150 hours at 50.degree. C., pH 5. At the end of the hydrolysis, glucose, which is water soluble, is in the liquid while unconverted cellulose, lignin, and other insoluble portions of the substrate remain in suspension. The glucose syrup is recovered by filtering the hydrolysis slurry; some washing of the fiber solids is carried out to increase the yield of glucose. The glucose syrup is then fermented to ethanol by yeast, and the ethanol recovered by distillation or other means. The ethanol fermentation and recovery are by well-established processes used in the alcohol industry.
The two-step process of pretreatment plus enzyme hydrolysis overcomes many of the problems associated with a single harsh acid hydrolysis. The specific action of the enzymes decreases the amount of degradation products and increases the yield of glucose. In addition, the fact that the pretreatment for fiber destruction is milder than that for cellulose hydrolysis means that lower chemical loadings can be used, decreasing the need for chemical recovery, and a lower amount of degradation products are made, increasing the yield and decreasing the inhibition of fermentation to ethanol by yeast.
Unfortunately, to date the approach of a pretreatment and an enzyme hydrolysis treatment has not been able to produce glucose at a sufficiently low cost, so as to make a fermentation to ethanol commercially attractive. Even with the most efficient currently known pretreatment processes, the amount of cellulase enzyme required to convert the cellulose to glucose is so high as to be cost-prohibitive for ethanol production purposes.
Several approaches have been taken to attempt to decrease the amount of cellulase enzyme required.
The approach of simply adding less cellulase to the system decreases the amount of glucose produced to an unacceptable extent.
The approach of decreasing the amount of enzyme required by increasing the length of time that the enzyme acts on the feedstock leads to uneconomical process productivity, stemming from the high cost of hydrolysis tanks.
The approach of reducing the amount of cellulase enzyme required by carrying out cellulose hydrolysis simultaneously with fermentation of the glucose by yeast is also inefficient. The so-called simultaneous saccharification and fermentation (SSF) process is not yet commercially viable because the optimum operating temperature for yeast, 28.degree. C., is too far below the optimum 50.degree. C. conditions required by cellulase. Operating a SSF system at a compromise temperature of 37.degree. C. is also inefficient, and invites microbial contamination.
The desire for a cost-effective ethanol production process has motivated a large amount of research into developing effective pretreatment systems. Such a pretreatment system would achieve all of the advantages of current pretreatments, including low production of degradation products and low requirements for chemical recovery, but with a sufficiently low requirement for cellulase enzymes so as to make the process economical.
The performance of a pretreatment system is characterized strictly by the amount of enzyme required to hydrolyze an amount of cellulose to glucose. Pretreatment A performs better than pretreatment B, if A requires less enzyme to produce a given yield of glucose than B.
The early work in pretreatment focused on the construction of a working device and determination of the conditions for the best performance.
One of the leading approaches to pretreatment is by steam explosion, using the process conditions described by FOODY (U.S. Pat. No. 4,461,648), which is incorporated herein by reference. In the FOODY process, biomass is loaded into a vessel known as a steam gun. Up to 1% acid is optionally added to the biomass in the steam gun or in a presoak. The steam gun is then filled very quickly with steam and held at high pressure for a set length of time, known as the cooking time. Once the cooking time elapses, the vessel is depressurized rapidly to expel the pretreated biomass, hence the terminology "steam explosion" and "steam gun".
In the FOODY process, the performance of the pretreatment depends on the cooking time, the cooking temperature, the concentration of acid used, and the particle size of the feedstock. The recommended pretreatment conditions in the FOODY process are similar for all the cellulosic feedstocks tested (hardwood, wheat straw, and bagasse) provided they are divided into fine particles. Furthermore, the cooking temperature is determined by the pressure of the saturated steam fed into the steam gun. Therefore, the practical operating variables that effect the performance of the pretreatment are the steam pressure, cooking time, and acid concentration. The FOODY process describes combinations of these variables for optimum performance; as one might expect, increasing the time decreases the temperature used, and vice versa. The range of steam pressure taught by FOODY is 250 psig to 1000 psig, which corresponds to temperatures of 208.degree. C. to 285.degree. C.
Another published study of steam explosion pretreatment parameters is Foody, et al, Final Report, Optimization of Steam Explosion Pretreatment, U.S. DEPARTMENT OF ENERGY REPORT ET230501 (April 1980). This study reported the effects of the pretreatment variables of temperature (steam pressure), particle size, moisture content, pre-conditioning, die configuration, and lignin content. The optimized steam explosion conditions were reported for three types of straws, five species of hardwood, and four crop residues.
The optimum pretreatment conditions as published by FOODY were subsequently confirmed by others using other feedstocks and different equipment. For example, GRETHLEIN (U.S. Pat. No. 4,237,226), describes pretreatment of oak, newsprint, poplar, and corn stover by a continuous plug-flow reactor, a device that is similar to an extruder. Rotating screws convey a feedstock slurry through a small orifice, where mechanical and chemical action break down the fibers.
GRETHLEIN specifies required orifice sizes, system pressures, temperatures (180.degree. C. to 300 C.), residence times (up to 5 minutes), acid concentrations (up to 1% sulfuric acid) and particle sizes (preferred 60 mesh). GRETHLEIN obtained similar results for all of the specified substrates he identified (See Column 3, line 30). Even though the GRETHLEIN device is quite different from the steam gun of FOODY, the time, temperature, and acid concentration for optimum performance are similar.
More recent work has focused on understanding the means by which pretreatment improves the enzymatic hydrolysis of a given substrate. BRINK (U.S. Pat. No. 5,628,830) describes the pretreatment of lignocellulosic material by using a steam process to break down the hemicellulose and following with hydrolysis of the cellulose using cellulase enzymes.
The first explanation offered to characterize the advantage of a pretreatment was that a pretreatment should be evaluated on the amount of lignin removed, with better performance associated with higher degrees of delignification. See Fan, Gharpuray, and Lee, Evaluation Of Pretreatments For Enzymatic Conversion Of Agricultural Residues, PROCEEDINGS OF THE THIRD SYMPOSIUM ON BIOTECHNOLOGY IN ENERGY PRODUCTION AND CONSERVATION, (Gatlinburg, Tenn., May 12-15, 1981). The notion that delignification alone characterizes pretreatment was also reported by Cunningham, et al, PROCEEDINGS OF THE SEVENTH SYMPOSIUM ON BIOTECHNOLOGY FOR FUELS AND CHEMICALS, (Gatlinburg, Tenn., May 14-17, 1985).
Grethlein and Converse, Common Aspects of Acid Prehydrolysis and Steam Explosion for Pretreating Wood, BIORESOURCE TECHNOLOGY 36(2):77-82 (1991), put forth the proposition that the degree of delignification is important only for previously dried substrates and, therefore, not a relevant consideration to most pretreatment processes that use undried feedstocks.
Knappert, et al, A Partial Acid Hydrolysis of Cellulosic Materials as a Pretreatment for Enzymatic Hydrolysis, BIOTECHNOLOGY AND BIOENGINEERING 23:1449-1463 (1980) reported that the increased susceptibility to enzyme hydrolysis after pretreatment is caused by the creation of micropores by the removal of the hemicellulose, a change in crystallinity of the substrate, and a gross reduction in the degree of polymerization of the cellulose molecule.
Grohmann, et al, Optimization of Dilute Acid Pretreatment of Biomass, SEVENTH SYMPOSIUM ON BIOTECHNOLOGY FOR FUELS AND CHEMICALS (Gatlinburg, Tenn., May 14-17, 1985) specifically supported one of the hypotheses of Knappert, et al by showing that removal of hemicellulose in pretreatment results in improved enzymatic hydrolysis of the feedstock. (See p. 59-80). Grohmann, et al worked with wheat straw and aspen wood at temperatures of 95.degree. C. to 160.degree. C. and cooking times of up to 21 hours. For both feedstocks, about 80% of the cellulose was digested by cellulase enzymes after optimum pretreatments, in which 80% to 90% of the xylan was removed from the initial material.
Grohmann and Converse also reported that the crystallinity index of the cellulose was not changed significantly by pretreatment. They further reported that pretreatments can create a wide range of degrees of polymerization while resulting in similar susceptibility to enzymatic hydrolysis.
Another alternative explanation offered for the improvements in enzymatic hydrolysis due to pretreatment is the increase in surface area of the substrate. Grethlein and Converse refined this explanation by showing that the surface area that is relevant is that which is accessible to the cellulase enzyme, which has a size of about 51 angstroms. The total surface area, which is measured by the accessibility of small molecules such as nitrogen, does not correlate with the rate of enzymatic hydrolysis of the substrate, for the reason that small pores that do not allow the enzyme to penetrate do not influence the rate of hydrolysis.
In spite of a good understanding of devices and optimum conditions for pretreatment, and a large quantity of research into the mechanism of a pretreatment process, there still does not exist an adequate pretreatment for a commercially feasible process to convert cellulosic materials to ethanol. Such a pretreatment process would be of enormous benefit in bringing the cellulose-to-ethanol process to commercial viability.